Wuschel2 enables highly efficient CRISPR/Cas-targeted genome editing during rapid de novo shoot regeneration in sorghum

• Published in: Communications biology Vol. 5; no. 1; p. 344
• Main Authors: Che, Ping, Wu, Emily, Simon, Marissa K., Anand, Ajith, Lowe, Keith, Gao, Huirong, Sigmund, Amy L., Yang, Meizhu, Albertsen, Marc C., Gordon-Kamm, William, Jones, Todd J.
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 11.04.2022; Nature Publishing Group; Nature Portfolio
• Subjects: 13; 13/106; 42; 42/41; 631/449; 631/61; Agrobacterium; Biology; Biomedical and Life Sciences; Callus; Cell culture; CRISPR; CRISPR-Cas Systems; Edible Grain - genetics; Gene Editing - methods; Genetic transformation; Genomes; Genotypes; Life Sciences; Plants, Genetically Modified - genetics; Regeneration - genetics; Sorghum; Sorghum - genetics; Sorghum bicolor; Tissue culture
• ISSN: 2399-3642; 2399-3642
• DOI: 10.1038/s42003-022-03308-w

For many important crops including sorghum, use of CRISPR/Cas technology is limited not only by the delivery of the gene-modification components into a plant cell, but also by the ability to regenerate a fertile plant from the engineered cell through tissue culture. Here, we report that Wuschel2 (Wus2) -enabled transformation increases not only the transformation efficiency, but also the CRISPR/Cas-targeted genome editing frequency in sorghum ( Sorghum bicolor L .). Using Agrobacterium -mediated transformation, we have demonstrated Wus2 -induced direct somatic embryo formation and regeneration, bypassing genotype-dependent callus formation and significantly shortening the tissue culture cycle time. This method also increased the regeneration capacity that resulted in higher transformation efficiency across different sorghum varieties. Subsequently, advanced excision systems and “altruistic” transformation technology have been developed to generate high-quality morphogenic gene-free and/or selectable marker-free sorghum events. Finally, we demonstrate up to 6.8-fold increase in CRISPR/Cas9-mediated gene dropout frequency using Wus2 -enabled transformation, compared to without Wus2 , across various targeted loci in different sorghum genotypes. Che et al. use Wuschel2-enabled genome transformation to induce somatic embryo formation in sorghum, a grain used in human food. Their approach not only overcomes the genotype-dependent barrier for genetic transformation without the introduction of morphogenic genes, but also increases the frequency of CRISPR/Castargeted genome editing.